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Efficient expression of a transgene in platelets using simian immunodeficiency virus-based vector harboring glycoprotein Ib promoter: in vivo model for platelet-targeting gene therapy

Tsukasa Ohmori^*, Jun Mimuro^*,, Katsuhiro Takano^*, Seiji Madoiwa^*,, Yuji Kashiwakura^*, Akira Ishiwata^*, Masanori Niimura^*, Katsuyuki Mitomo, Toshiaki Tabata, Mamoru Hasegawa, Keiya Ozawa^, and Yoichi Sakata^*,,1

^* Research Division of Cell and Molecular Medicine, Center for Molecular Medicine, Jichi Medical School, Tochigi, Japan;

Hematology Division of Department of Medicine, Jichi Medical School, Tochigi, Japan;

DNAVEC Corp., Ibaraki Japan; and

Research Division of Genetic Therapeutics, Center for Molecular Medicine, Jichi Medical School, Tochigi, Japan

¹Correspondence: Research Division of Cell and Molecular Medicine, Center for Molecular Medicine, Jichi Medical School, Minamikawachi, Tochigi 329-0498, Japan. E-mail: yoisaka{at}jichi.ac.jp

ABSTRACT

Platelets release several mediators that modify vascular integrity and hemostasis. In the present study, we developed a technique for efficient transgene expression in platelets in vivo and examined whether this targeted-gene-product delivery system using a platelet release reaction could be exploited for clinical applications. Analysis of luciferase reporter gene constructs driven by platelet-specific promoters (the GPIIb, GPIb, and GPVI) revealed that the GPIb promoter was the most potent in the megakaryoblastic cell line UT-7/TPO and human CD34⁺-derived megakaryocytes. Transduction of UT-7/TPO; CD34⁺-derived megakaryocytes; and c-Kit⁺, ScaI⁺, and Lineage^– (KSL) murine hematopoietic stem cells with a simian immunodeficiency virus (SIV)-based lentiviral vector carrying eGFP resulted in efficient, dose-dependent expression of eGFP, and the GPIb promoter seemed to bestow megakaryocytic-specific expression. Transplantation of KSL cells transduced with SIV vector containing eGFP into mice showed that there was preferable expression of eGFP in platelets driven by the GPIb promoter [7–11% for the cytomeglovirus (CMV) promoter, 16–27% for the GPIb promoter]. Furthermore, transplantation of ex vivo-transduced KSL cells by SIV vector carrying human factorVIII (hFVIII) driven by the GPIb promoter induced the production of detectable transcripts of the hFVIII gene and the hFVIII antigen in bone marrow and spleen for at least 90 days and partially corrected the hemophilia A phenotype. Platelet-targeting gene therapy using SIV vectors appears to be promising for gene therapy approaches toward not only inherited platelet diseases but also other hemorrhagic disorders such as hemophilia A.—Ohmori, T., Mimuro, J., Takano, K., Madoiwa, S., Kashiwakura, Y., Ishiwata, A., Niimura, M., Mitomo, K., Tabata, T., Hasegawa, M., Ozawa, K., Sakata, Y. Efficient expression of a transgene in platelets using simian immunodeficiency virus-based vector harboring glycoprotein Ib promoter: in vivo model for platelet-targeting gene therapy.

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